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Subepithelial myofibroblasts enhance stem cell regeneration after intestinal epithelial damage [scRNA-Seq]

ABSTRACT: The intestinal epithelium relies on the surrounding niche environment to maintain homeostasis, but how the niche supports intestinal stem cell regeneration is unclear. We analyzed a number of distinct mesenchymal populations in the intestinal stem cell niche following two models of murine intestinal injury, including whole body irradiation and diphtheria-toxin injected Lgr5Dtr mice. We isolated Pdgfra fibroblasts from the murine transgenic PDGFRAH2BEGFP (GFP-hi and GFP-lo) and PdgfraCreERT2;Rosa26tDtomato (tDtomato+) mice, smooth muscle from Myh11CreERT2;PdgfraH2BeGFP;Rosa26tDtomato (GFP-tDtomato+), and blood and lympathic endothelium via antibody for CD31+LYVE1- and CD31+LYVE1+, respectively. We found distinct molecular signatures of these cell types following stem cell loss and focused our analyses on known trophic factors of the overlaying intestinal epithelium.

ORGANISM(S): Mus musculus

PROVIDER: GSE322565 | GEO | 2026/09/18

REPOSITORIES: GEO

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Subepithelial myofibroblasts enhance stem cell regeneration after intestinal epithelial damage [RNA-Seq]

Project description:The intestinal epithelium relies on the surrounding niche environment to maintain homeostasis, but how the niche supports intestinal stem cell regeneration is unclear. We analyzed a number of distinct mesenchymal populations in the intestinal stem cell niche following two models of murine intestinal injury, including whole body irradiation and diphtheria-toxin injected Lgr5Dtr mice. We isolated Pdgfra fibroblasts from the murine transgenic PDGFRAH2BEGFP (GFP-hi and GFP-lo) and PdgfraCreERT2;Rosa26tDtomato (tDtomato+) mice, smooth muscle from Myh11CreERT2;PdgfraH2BeGFP;Rosa26tDtomato (GFP-tDtomato+), and blood and lympathic endothelium via antibody for CD31+LYVE1- and CD31+LYVE1+, respectively. We found distinct molecular signatures of these cell types following stem cell loss and focused our analyses on known trophic factors of the overlaying intestinal epithelium.

2026-09-18 | GSE322564 | GEO

Distinct mesenchymal cell populations generate the essential intestinal BMP signaling gradient

Project description:The intestinal epithelium requires trophic factors and signaling gradients to maintain proper positioning of Intestinal Stem Cells (ISC) and proper differentiation of ISCs. Because the underlying mesenchyme is composed of numerous cell types, we utilized both transgenic and antibody methods to isolate specific mesenchymal populations including GFP-hi and GFP-lo cells from the murine transgenic PDGFRAH2BEGFP mouse (Jackson, #007669), endothelial and lymphatic endothelial cells via antibody for CD31+LYVE1- and CD31+LYVE1+, respectively. We found distinct molecular signatures of these cell types and focused our analyses on known trophic factors of the overlaying intestinal epithelium.

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Project description:Intestinal stem cells are sequestered into crypts generated during postnatal development in mice. During this timeframe, we investigated the transition of the surrounding mesenchymal niche. Using single cell RNA sequencing, we generated transcriptomic libraries from a combination of whole tissue and transgenically labeled fractions, including GFP+ fractions from PdgfraH2BEGFP and GFP-tDt+ fractions from PdgfraH2BEGFP;Myh11CreERT2;Rosa26tDt+ mice, of the intestinal mesenchyme at varying postnatal and adult stages. We found distinct molecular signatures of these cell types, focusing our analysis on the temporal changes and known epithelial trophic factors.

2023-02-08 | GSE184158 | GEO

Bulk RNA sequencing of endothelial-derived and non-endothelial-derived BMSCs and OLCs of VE-Cadherin-Cre;LoxP-tdTomato mice

Project description:We performed lineage tracing experiments using VE-Cadherin-Cre;LoxP-tdTomato mice. In these mice, endothelial cells (ECs) and their progeny are permanently marked by tdTomato fluorescence. We found that a substantial subset of stromal cells is derived from ECs, as indicated by their tdTomato expression. These findings support the notion that endothelial to mesenchymal transition (EndoMT) contributes to hematopoietic bone marrow niche formation in mice. Here we sought to determine the transcriptomic differences between endothelial-derived (tdTomato-positive) and non-endothelial-derived (tdTomato-negative) bone marrow stromal cells (BMSCs) and osteo/chondrolineage progenitor cells (OLCs). Murine niche populations were obtained from collagenased bone fraction of VE-Cadherin-Cre;LoxP-tdTomato mice at 3 weeks (n=2) or 11 weeks (n=2) of age. BMSCs (CD45-TER119-CD31-CD144-SCA-1+ CD51+ cells) and OLCs (CD45-TER119-CD31-CD144-Sca1-CD51+ cells) were FACS-purified and sequenced.

2021-01-25 | E-MTAB-10020 | biostudies-arrayexpress

Intestinal subepithelial myofibroblasts support the growth of intestinal epithelial stem cells

Project description:Intestinal epithelial stem cells (ISCs) are the focus of recent intense study. Current in vitro models rely on supplementation with the Wnt agonist R-spondin1 to support robust growth, ISC self-renewal, and differentiation. Intestinal subepithelial myofibroblasts (ISEMFs) are important supportive cells within the ISC niche. We hypothesized that co-culture with ISEMF enhances the growth of ISCs in vitro and allows for their successful in vivo implantation and engraftment. ISC-containing small intestinal crypts, FACS-sorted single ISCs, and ISEMFs were procured from C57BL/6 mice. Crypts and single ISCs were grown in vitro into enteroids, in the presence or absence of ISEMFs. ISEMFs enhanced the growth of intestinal epithelium in vitro in a proximity-dependent fashion, with co-cultures giving rise to larger enteroids than monocultures. Co-culture of ISCs with supportive ISEMFs relinquished the requirement of exogenous R-spondin1 to sustain long-term growth and differentiation of ISCs. Mono- and co-cultures were implanted subcutaneously in syngeneic mice. Co-culture with ISEMFs proved necessary for successful in vivo engraftment and proliferation of enteroids; implants without ISEMFs did not survive. ISEMF whole transcriptome sequencing and qPCR demonstrated high expression of specific R-spondins, well-described Wnt agonists that supports ISC growth. Specific non-supportive ISEMF populations had reduced expression of R-spondins. The addition of ISEMFs in intestinal epithelial culture therefore recapitulates a critical element of the intestinal stem cell niche and allows for its experimental interrogation and biodesign-driven manipulation. Two samples of intestinal subepithelial myofibroblasts were used in this study.

2013-12-20 | E-GEOD-52402 | biostudies-arrayexpress

Analysis of thymic dendritic cells (DCs) subtypes from Foxn1cre x ROSA26TdTOMATO mice by single cell transcription profiling.

Project description:Thymi from Foxn1cre x ROSA26TdTOMATO were enzymaticaly digested, cells were MACS enriched for CD11c+ fraction. Dendritic cells were then FACS sorted (BD influx sorter) as Gr-1-CD11c+TdTOMATO+. TdTOMATO DCs represent fraction of thymic DCs which acquire antigen (TdTOMATO) from thymic epithelial cells, through the process of antigen transfer.

2020-03-20 | E-MTAB-8028 | biostudies-arrayexpress

Intestinal subepithelial myofibroblasts support the growth of intestinal epithelial stem cells

2013-12-20 | GSE52402 | GEO

Distinct populations of embryonic epithelial progenitors generate Lgr5+ intestinal stem cells

Project description:To follow the changes in the transcriptional programs accompanying the specification of the embryonic Lgr5+ cells we sequenced whole transcriptomes of embryonic intestinal epithelium progenitors (at E13.5 and E15.5). EpCAM positive embryonic gut epithelium was isolated from dissected small intestines using fluorescence activated cell sorting (FACS). Lgr5+ progenitors were purified on the basis of GFP fluorescence from Lgr5GFP-Cre-ERT mice (Barker et al. 2007). Double positive embryonic progenitors were isolated by FACS based on GFP and tdTomato fluorescence.

2019-09-24 | GSE104727 | GEO

Bulk RNA-Sequencing of mouse large intestinal lymphatics (capillaries and collectors)

Project description:Purpose: Profile global gene expression of intestinal lymphatics to compare with published datasets Methods: Intestinal tissues were digested, lymphatic endothelial cells were sorted using lymphatic markers including CD31, Podoplanin, Lyve1. Total RNA was isolated from sorted cells using the Direct-Zol RNA MiniPrep kit (Zymo). Samples were submitted for bulk RNA-Sequencing. Conclusions: Gobal gene expression profile of intestinal lymphatics for further comparison with published datasets

2022-06-21 | GSE199082 | GEO

Id2 controls specification of Lgr5+ intestinal stem cell progenitors during gut development

Project description:To follow the changes in the transcriptional programs accompanying the specification of the adult ISCs we sequenced whole transcriptomes of embryonic intestinal epithelium progenitors (at E11.5 and E12.5) and adult ISCs. EpCAM positive embryonic gut epithelium was isolated from dissected small intestines using fluorescence activated cell sorting (FACS). Adult ISCs were purified on the basis of GFP fluorescence from crypts of Lgr5GFP-Cre-ERT mice (Barker et al. 2007) Double positive adlut ISCs were isolated by FACS based on GFP and tdTomato fluorescence.

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